Abstract: Exposure of healthy wheat seeds (Triticum aestivum var Sonalika) to mild dose of cadmium (Cd2+) given as 50 uM CdCl2 for 48 h and then washed off Cd2+ offered resistance to the subsequent infection by Fusarium oxysporum inoculum. Seven days old seedlings having two primary leaves were aseptically inoculated with fungus, F. oxysporum (1 × 106) spores. The seedlings pre-exposed to low level of Cd2+ survived the Fusarium infection, while plantlets without Cd2+ stress wilted and then perished due to Fusarium infection. The stress associated proteins induced by Cd2+ (50 u), F. oxysporum and by the co-stress (50 uM Cd2+ and then with F. oxysporum) treatments were observed to be of same molecular weight (51 kDa). Antibody was raised against the purified Cd2+-stress associated protein (CSAP). Immuno-gold labeling of wheat seedling root tissue showed the presence of this CSAP in Cd2+ pre-exposed and in co-stressed tissues and to be located predominantly on the inner linings of the cell membranes. We also observed that the anti-CSAP-antibody also labeled the root tissue of only Fusarium inoculated seedlings and the gold labeling was intensely located on the membrane. This cross-reaction of anti-CSAP suggests that Fusarium-induced stress protein (FISP) possibly has close homology to CSAP. We thus show for the first time the over expression of a high molecular mass protein by mild dose of Cd2+ pre-exposure to wheat seeds which subsequently provided protection against Fusarium infection. This mode of resistance developed by an abiotic stress-causing agent against pathogen infection is novel.
Abstract: Guaiacol-peroxidase (POX) and polyphenol oxidase (PPO) activities were measured spectrophotometrically in resistant (cvs. Sumai#3 and Wang shui-bai) and susceptible (cvs. Falat and Golestan) wheat heads at flowering, milk, dough and ripening stages following the inoculation with Fusarium graminearum at anthesis. POX specific activity in resistant and susceptible wheat cultivars showed a significant increase during the milk stage as compared with the non-inoculated control plants. POX activity reached the highest level in heads of Wang shui-bai followed by those of Falat, Sumai#3 and Golestan cultivars at milk stage. The optimal pH for POX activity was 5.4. POX was inhibited by KCN. Native polyacrylamide gel revealed the presence of upto three basic (cationic) and six acidic (anionic) isozymes in wheat heads after a specific activity stain. PPO specific activity in wheat heads reached a maximum level during the milk stage and subsequently declined. This activity was three times higher in the resistant cultivars than the non-inoculated control plants. In Falat and Golestan cultivars, PPO activity level was half of those in resistant cultivars. The optimal pH for PPO was 6.4. PPO-catalyzed reaction was inhibited by ascorbic acid. Activity stain in non-denaturing polyacrylamide gel revealed the presence of one basic and six acidic isozymes in wheat heads. The susceptible Falat heads pre-treated with an autoclaved mycelial wall preparation showed induced resistance against FHB and increased activities of POX and PPO.
Abstract: Defense responses to inoculation with Fusarium oxysporum Schlecht f. sp. lupini were studied in embryo axes of Lupinus luteus L. cv. Polo cultured on a medium with sucrose (60 mM) or without it. Exogenous sucrose caused a marked endogenous increase in concentrations of sucrose, glucose and fructose in embryo axes. In axes cultured with sucrose, high performance liquid chromatography (HPLC) revealed generally higher levels of isoflavone glycosides (particularly until 48 h of culture) and free aglycones (genistein, wighteone, luteone). Inoculation resulted in a considerable decline in soluble carbohydrates between 24 and 72 h of culture. Simultaneously, the infection stimulated an increase in the level of free isoflavone aglycones in inoculated embryo axes, as compared to non-inoculated ones. Concentrations of free aglycones (i.e. genistein, wighteone and luteone) after infection were particularly high in inoculated embryo axes fed with sucrose. Genistein was a better inhibitor to F. oxysporum growth than genistein 7-O-glucoside tested. Exogenous sucrose also stimulated the activity of phenylalanine ammonialyase (PAL, EC 4.3.1.5)—an important enzyme initiating phenylpropanoid metabolism. After infection of tissues, a strong increase was observed in the activity of PAL and b-glucosidase (EC 3.2.1.21)—an enzyme hydrolyzing isoflavone glycosides. Furthermore, the growth of inoculated embryo axes cultured with sucrose was less inhibited as a result of infection than inoculated axes cultured under carbohydrate deficiency conditions. Additionally, it had been reported previously that disease symptoms of embryo axes growing in the presence of sucrose were less intensive [30]. These results suggest that soluble sugars are involved in the mechanism of resistance, as they can stimulate phenylpropanoid metabolism and contribute to the increase in concentration of isoflavonoids, which are important elements of the defense system of legumes.
Abstract: Peroxidase activity (EC 1.11.1.7) towards phenolic substrates, i.e. pyrogallol, syringaldazine and guaiacol, and ascorbate peroxidase activity (EC 1.11.1.11) were analyzed in embryo axes of Lupinus luteus L. cv. Polo cultured on Heller medium for 96 h after inoculation with the necrotrophic fungus Fusarium oxysporum f.sp. Schlecht lupini. Four variants were compared: inoculated embryo axes cultured with 60 mM sucrose (+Si) or without it (-Si), and non-inoculated embryo axes cultured with 60 mM sucrose (+Sn) or without it (-Sn). Between 0 and 96 h of culture, peroxidase activity towards the phenolic substrates increased in all variants except -Si, where a decrease was noted in peroxidase activity towards syringaldazine and guaiacol, but not towards pyrogallol. In +Si tissues, a considerable increase in enzyme activity towards these substrates was recorded starting from 72 h of culture. Lignin content of +Si tissues increased already at the first stage of infection, i.e. 24 h after inoculation. Additionally, in +Sn tissues, high ascorbate peroxidase activity was observed during the culture. Its activity increased in +Si tissues, beginning at 72 h after inoculation. However, this was lower than in +Sn tissues. At 72 h after inoculation, a considerably stronger development of the infection was observed in -Si than in +Si tissues during our earlier research [Morkunas, I. et al., 2005. Sucrose-stimulated accumulation of isoflavonoids as a defense response of lupine to Fusarium oxysporum. Plant Physiol Biochem 2005; 43: 363–73]. Both peroxidases assayed towards phenolic substrates and ascorbate peroxidase was less active in -Si tissues than in -Sn tissues. Hydrogen peroxide concentration was much higher in -Si than in +Si tissues. These results indicate that peroxidases may be some of the elements of the defense system that are stimulated by sucrose in yellow lupine embryo axes in response to infection caused by F. oxysporum.
Abstract: A few EST-derived STS markers localized on Qfhs.ndsu-3BS, a major QTL for resistance to Fusarium head blight (FHB) in wheat,have been previously identified in the Sumai 3/Stoa population. In this study, we used a Wangshuibai(resistant)/Seri82 (susceptible)derived population, linkage group, QTL, and quantitative gene expression analysis to assess the genetic background dependence and stability of the EST-derived STS markers for use in marker aided selection to improve FHB resistance in wheat. Based on our results, a QTL in the map interval of Xsts3B-138_1-Xgwm493 on chromosome 3BS was detected for FHB resistance, which accounted for up to 16% of the phenotypic variation. BLASTN analysis indicated that Xsts3B-138_1 sequence had significant similarity with the resistance gene analogue. Real-time quantitative PCR showed that the relative expression of Xsts3B-138_1 in Wangshuibai at 96 h after inoculation was 2.6 times higher than Seri82. Our results underlined that EST-derived STS3B-138 markers could be predominantly used in marker aided selection to improve FHB resistance in wheat.
Abstract: The in vitro antifungal properties of chitosan and its role in protection of soybean from a sudden death syndrome (SDS) were evaluated. Chitosan inhibited the radial and submerged growth of F. solani f. sp. glycines with a marked effect at concentrations up to 1mg/ml indicating antifungal property and at 3mg/ml was able to delay SDS symptoms expression on soybean leaves for over three days after fungal inoculation when applied preventively. Chitosan was able to induce the level of chitinase activity in soybean resulting in the retardation of SDS development in soybean leaves. However, the SDS symptoms gradually appeared and were associated with the reduction of chitinase activity level after five days of infection period. These results suggested the role of chitosan in partially protecting soybeans from F. solani f. sp. glycines infection.
Abstract:Clonostachys rosea (CR) is a common worldwide saprophyte with destructive effect against several plant pathogenic fungi showing antagonistic features against a wide variety of pathogens. We recently isolated a strain of C. rosea, named CR47, from wheat crown infected with Fusarium culmorum (FC); this strain proved to be effective against Fusarium seed borne diseases of cereals under field condition. In this paper the function of C. rosea applied as seed treatment on wheat seedling growth was investigated. In addition, we investigated the expression pattern of peroxidases and chitinases as well as PR4 proteins following both CR treatments of seeds and FC infection and also in the three-component system pathogen–antagonist–wheat. Several chitinase isoforms were induced by CR-treatment both in coleoptiles and roots, whereas some peroxidase isoforms were induced only in the presence of both antagonist and pathogen. In the latter case, it seems that CR-treatment by itself promotes plant growth and reduces the peroxidase expression, while enhances some chitinase isoforms probably involved in cell wall disruption. Moreover, both the antagonist and the pathogen studied induced PR4 protein expression, which probably exerts its role on the invading microorganisms by a translation-inhibitory process that could be ascribed to their ribonuclease activity.
Abstract: Selected isolates of Pseudomonas fluorescens (Pf1–94, Pf4–92, Pf12–94, Pf151–94 and Pf179–94) and chemical resistance inducers (salicylic acid, acetylsalicylic acid, DL-norvaline, indole-3-carbinol and lichenan) were examined for growth promotion and induced systemic resistance against Fusarium wilt of chickpea. Amarked increase in shoot and root length was observed in P. fluorescens treated plants. The isolatesof P. fluorescens systemically induced resistance against Fusarium wilt of chickpea caused by Fusarium. oxysporum f.sp. ciceri (FocRs1), and significantly (P = 0.05) reduced the wilt disease by 26–50% as compared to control. Varied degree of protection against Fusarium wilt was recorded with chemical inducers. The reduction in disease was more pronounced when chemical inducers were applied with P. fluorescens. Among chemical inducers, SA showed the highest protection of chickpea seedlings against wilting. Fifty two- to 64% reduction of wilting was observed in soil treated with isolate Pf4–92 along with chemical inducers. A significant (P = 0.05; r = –0.946)negative correlation was observed in concentration of salicylic acid and mycelial growth of FocRs1 and at a concentration of 2000 µg ml–1 mycelial growth was completely arrested. Exogenously supplied SA also stimulated systemic resistance against wilt and reduced the disease severity by 23% and 43% in the plants treated with 40 and 80 µg ml–1 of SA through root application. All the isolates of P. fluorescens produced SA in synthetic medium and in root tissues. HPLC analysis indicated that Pf4–92 produced comparatively more SA than the other isolates. 1700 to 2000 ug SA g–1 fresh root was detected from the application site of root after one day of bacterization whereas, the amount of SA at distant site ranged between 400–500 ug. After three days of bacterization the SA level decreased and was found more or less equal at both the detection sites.
Abstract:Fusarium verticillioides causes maize ear rot and contaminates kernels with fumonisin mycotoxins in Argentina. The aim of this work was to elucidate if the kernel pericarp and its surface wax layer are resistance factors to fumonisin accumulation in maize genotypes from Argentina. Fourteen maize genotypes were inoculated with F. verticillioides in laboratory assays. Intact kernels of genotypes resistant to fumonisin accumulation in the field had the lowest mycotoxin concentration in the current assays suggesting that kernel factors are involved. Intact kernels of landraces, breeding populations and L4637 inbreed were less susceptible than wounded ones, suggesting that intact kernel pericarp restricted fumonisin accumulation. Removing wax from the pericarp significantly increased fumonisin concentration and a higher wax content on kernels was associated to lower fumonisin accumulation. Our results suggest that the pericarp and its wax content are resistance factors to fumonisin accumulation in most genotypes assayed. Nevertheless, other kernel factors could not be excluded.
Abstract: Overexpression (OE) was used to study the role of the Arabidopsis Golden2-like (GLK1) transcriptional activator in regulating gene expression. Affymetrix Gene Chip and RT-PCR analyses indicated that GLK1 OE in Arabidopsis reprogrammed gene expression networks to enhance a high constitutive expression of genes encoding disease defense related proteins. These include PR10, isochorismate synthase, antimicrobial peptides, glycosyl hydrolases, MATE efflux and other genes associated with pathogen response and detoxification. However, PR1, an indicator of systemic acquired resistance (SAR), was down regulated in GLK1 OE. GLK1 OE in Arabidopsis confers resistance to Fusarium graminearum, a broad host pathogen responsible for major losses in cereal crops. This is the first identification of the GLK1 regulon and a novel role for GLK1 in plant defense, suggesting its potential use for providing disease resistance in crop plants.
Abstract